A region N-terminal to the tandem SH3 domain of p47phox plays a crucial role in the activation of the phagocyte NADPH oxidase
A region N-terminal to the tandem SH3 domain of p47phox plays a crucial role in the activation of the phagocyte NADPH oxidase
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DOI：
10.1042/bj20082028
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发表时间：
2009-04-15
影响因子：
4.1
通讯作者：
Sumimoto, Hideki
中科院分区：
文献类型：
--
作者：
Taura, Masahiko;Miyano, Kei;Sumimoto, Hideki
吞噬细胞中产生超氧化物的NADPH氧化酶对宿主防御至关重要;其催化核心是膜整合蛋白gp 91（phox）[也称为Nox 2（NADPH氧化酶2）]，它与p22 P（phox）形成稳定的异源二聚体。氧化酶的激活需要三种胞质蛋白p47（phox）、p67（phox）和小GT3 Rac的膜转位。在膜上，这些蛋白质与gp 91（phox）-p22（phox）异二聚体组装，并诱导gp 91（phox）的构象变化，导致超氧化物的产生。p47（phox）利用其两个串联排列的SH 3结构域易位到膜上，SH 3结构域直接与P22（phox）相互作用，而P67（phox）以P47（phox）依赖性方式募集。在本研究中，我们表明，一个短的区域N-末端的双SH 3结构域是需要激活的吞噬细胞NADPH氧化酶。丙氨酸取代IIe（152）在这个区域，是一个残基，在进化过程中是完全保守的，导致失去激活氧化酶的能力;和Thr（153）的替代也阻止氧化酶的激活，但在较小程度上。此外，p47（phox）同源物Noxol（Nox组织者1）的相应异亮氨酸残基（IIe（151））参与非吞噬性氧化酶（如Nox 1和Nox 3）的活化。p47（phox）中的I152 A取代。然而，不影响其与P22（phox）或与p67（phox）的相互作用。与此一致，突变体P47（phox）（I152 A）以及野生型蛋白质在细胞刺激时靶向膜，并且P67（phox）和Rac的膜募集通常发生在表达P47（phox）（I152 A）的细胞中。因此，p47（phox）的IIe（152）区在氧化酶激活中起着至关重要的作用，可能是通过在氧化酶组装后的过程中发挥作用。
The superoxide-producing NADPH oxidase in phagocytes is crucial for host defence; its catalytic core is the membrane-integrated protein gp91(phox) [also known as Nox2 (NADPH oxidase 2)], which forms a stable heterodimer with p22P(phox). Activation of the oxidase requires membrane translocation of the three cytosolic proteins p47(phox), p67(phox) and the small GTPase Rac. At the membrane, these proteins assemble with the gp91(phox)-p22(phox) heterodimer and induce a conformational change of gp91(phox), leading to Superoxide production. p47(phox) translocates to membranes using its two tandemly arranged SH3 domains, which directly interact with P22(phox), whereas P67(phox) is recruited in a P47(phox)-dependent manner. In the present study, we show that a short region N-terminal to the bis-SH3 domain is required for activation of the phagocyte NADPH oxidase. Alanine substitution for IIe(152) in this region, a residue that is completely conserved during evolution, results in a loss of the ability to activate the oxidase; and the replacement of Thr(153) also prevents oxidase activation, but to a lesser extent. In addition, the corresponding isoleucine residue (IIe(151)) of the p47(phox) homologue Noxol (Nox organizer 1) participates in the activation of non-phagocytic oxidases, such as Nox1 and Nox3. The I152A substitution in p47(phox). however, does not affect its interaction with P22(phox) or with p67(phox). Consistent with this, a mutant P47(phox) (I152A), as well as the wild-type protein, is targeted upon cell stimulation to membranes, and membrane recruitment of P67(phox) and Rac normally occurs in P47(phox) (I152A)-expressing cells. Thus the IIe(152)-containing region of p47(phox) plays a crucial role in oxidase activation, probably by functioning at a process after oxidase assembly.